ABSTRACT
Apoptotic vesicles (apoVs) are apoptotic-cell-derived nanosized vesicles that take on dominant roles in regulating bone homeostasis. We have demonstrated that mesenchymal stem cell (MSC)-derived apoVs are promising therapeutic agents for bone regeneration. However, clinical translation of MSC-derived apoVs has been hindered due to cell expansion and nuclear substance. As another appealing source for apoV therapy, blood cells could potentially eliminate these limitations. However, whether blood cells can release apoVs during apoptosis is uncertain, and the detailed characteristics and biological properties of respective apoVs are not elucidated. In this study, we showed that platelets (PLTs) could rapidly release abundant apoVs during apoptosis in a short time. To recognize the different protein expressions between PLT-derived apoVs and PLTs, we established their precise protein landscape. Furthermore, we identified six proteins specifically enriched in PLT-derived apoVs, which could be considered as specific biomarkers. More importantly, PLT-derived apoVs promoted osteogenesis of MSCs and rescued bone loss via Golgi phosphoprotein 2 (GOLPH2)-induced AKT phosphorylation, therefore, leading to the emergence of their potential in bone regeneration. In summary, we comprehensively determined characteristics of PLT-derived apoVs and confirmed their roles in bone metabolism through previously unrecognized GOPLH2-dependent AKT signaling, providing more understanding for exploring apoV-based therapy in bone tissue engineering.
Subject(s)
Mesenchymal Stem Cells , Proto-Oncogene Proteins c-akt , Proto-Oncogene Proteins c-akt/metabolism , Mesenchymal Stem Cells/metabolism , Bone Regeneration , Blood Platelets/metabolism , Osteogenesis , Phosphoproteins/metabolismABSTRACT
Golgi phosphoprotein 2 (GP73) is highly expressed in hepatocellular carcinoma (HCC) cells, where it serves as a biomarker and indicator of disease progression. We used MTS assays, anchorage-independent cell colony formation assays and a xenograft tumor model to show that GP73-specific siRNAs inhibit HCC proliferation in HepG2, SMMC-7721, and Huh7 cell lines and in vivo. Following GP73 silencing, levels of p-Rb, a factor related to metastasis, were reduced, but cell cycle progression was unaffected. Our results suggest that GP73 silencing may not directly suppress proliferation, but may instead inhibit cell motility. Results from proliferation assays suggest GP73 reduces expression of epithelial mesenchymal transition (EMT)-related factors and promotes cell motility, while transwell migration and invasion assays indicated a possible role in metastasis. Immunofluorescence co-localization microscopy and immunoblotting showed that GP73 decreases expression of N-cadherin and E-cadherin, two key factors in EMT, which may in turn decrease intracellular adhesive forces and promote cell motility. This study confirmed that GP73 expression leads to increased expression of EMT-related proteins and that GP73 silencing reduces HCC cell migration in vitro. These findings suggest that GP73 silencing through siRNA delivery may provide a novel low-toxicity therapy for the inhibition of tumor proliferation and metastasis.
Subject(s)
Carcinoma, Hepatocellular/genetics , Liver Neoplasms/genetics , Membrane Proteins/genetics , RNA Interference , Animals , Cadherins/metabolism , Carcinoma, Hepatocellular/metabolism , Carcinoma, Hepatocellular/pathology , Cell Line , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Hep G2 Cells , Humans , Immunoblotting , Liver Neoplasms/metabolism , Liver Neoplasms/pathology , Male , Membrane Proteins/metabolism , Mice, Inbred BALB C , Mice, Nude , Microscopy, Confocal , Reverse Transcriptase Polymerase Chain Reaction , Transplantation, HeterologousABSTRACT
As a novel Golgi protein,GOLPH2 expression is associated with maintaining the structural integrity of the Golgi apparatus.As an oncoprotein,GOLPH2 expression is associated with malignant biological behavior of cancer cells.Important roles of GOLPH2 in the tumorigenesis and metastasis make it become a new target for tumor diagnosis and treatment.
